The permissive role played by the enteric microenvironment in the colonization of the bowel and its influence on the phenotypic expression of the neural crest-derived precursors of the neurons and glia of the mammalian enteric nervous system (ENS) will be analyzed. Experiments will focus on the lethal spotted (ls/ls) mutant mouse, an animal in which the terminal portion of the gut is congenitally aganglionic. We have obtained evidence that this aganglionosis is due to a localized abnormality of the non-neuronal tissue of the gut, which fails to support the ingrowth and/or survival of the precursors of intrinsic enteric neurons. The nature of the defect will now be analyzed in vitro. Chimeric cultures will be used so that neural crest cells and their progeny can be identified. Effects on enteric neuronal and glial development of elements of the ls/ls enteric mesenchyme, including soluble factors, extracellular matrix, and cell to cell contact will be assessed. In addition, grafts of murine gut will be placed into the vagal neural crest migration pathway of chick embryos, both to determine if the presumptive aganglionic ls/ls gut can be colonized by normally migrating crest cells, and to determine if neural crest migratory pathways can provide a route of escape for neural precursors from the presumptive aganglionic tissue. A final series of experiments will evaluate the role of cell division in the development of enteric neurons. We will test the hypothesis that after the first enteric neurons appear the gut still retains a pool of proliferating neuroblasts from which peptidergic neurons are derived.